Clemson researchers receive $1.8 million for root study with broad implications for agriculture

CLEMSON – Three Clemson University scientists have been awarded a $1.8 million grant to study gene expression in roots that eventually could pave the way for increased crop production, save the world billions of dollars in fertilizer usage and help rid the planet of a damaging pollutant.

Julia Frugoli, Alex Feltus and Victoria Corbin are the recipients of the three-year National Science Foundation grant. Their project will focus on legumes (such as peas and beans), which are the only crops capable of teaming with bacteria in their roots to chemically convert nitrogen from the air we breathe into a form that living organisms can use. All other crops can only obtain this essential nutrient from soil. Nitrogen gas makes up more than 70 percent of the atmosphere, but it is inert and therefore useless to plants and animals until it is converted or "fixed."

"Legumes have a special relationship with soil bacteria that know how to take nitrogen out of the air," said Frugoli, Alumni Distinguished Professor and associate chair of genetics and biochemistry at Clemson. "The plants make nodules in their roots for the bacteria to live in. These bacteria convert the nitrogen and feed it to the plant. In return, the plant feeds the bacteria carbon. This makes it possible for legumes to grow healthy and strong even when no nitrogen fertilizer is added to depleted soils."

In 2011, the U.S. Department of Agriculture reported that 12.8 billion tons of nitrogen fertilizer costing more than $800 million were added to fields that produce corn, cotton, grains and other commodity crops. Yet only about half of this nitrogen is taken up by crops. The rest enters the water table, polluting local wells and running off into rivers, lakes and oceans, overloading them with nitrates that create "dead zones" in which aquatic life cannot survive. Fertilizers also release nitrous oxide, a greenhouse gas with 300 times the heat-trapping capacity of carbon dioxide. Meanwhile, in developing nations where synthetic fertilizers are unaffordable, nitrogen is often the limiting nutrient for growing crops, leading to poor yields.

"If all plants could do what legumes can do, then we wouldn't need nitrogen fertilizer and we would solve these problems," said Frugoli, whose lab at Clemson works to understand the signaling between the plant and bacteria in the nitrogen fixation relationship. "So the goal of this project is to fully understand – at the cellular and molecular level – how and why legumes are able to create this symbiotic relationship with bacteria. This would be a first step to establishing nitrogen-fixing symbioses in crops that do not form these kinds of root nodules."

While Frugoli is known for her research on the legume plant system, Feltus is equally renowned for his expertise in bioformatics, which is the science of collecting and analyzing complex biological data locked into DNA genetic code. In this project, Frugoli's role is to extract ribonucleic acid (RNA) from individual cells in plant tissue. The RNA is then turned into deoxyribonucleic acid (DNA), which can be sequenced using the latest available technologies, then analyzed on the Palmetto Cluster supercomputer at Clemson. RNA is present in all living cells and acts as a messenger for DNA, which is the carrier of genetic information.

"I will couple advanced DNA sequencing and computer technologies to 'see' gene expression, and I will provide Dr. Frugoli with the potential genes that are enabling symbiosis based upon where and how they are expressed in the root," said Feltus, associate professor in genetics and biochemistry and director of the Clemson Systems Genetics Lab. "I'll be finding the holistic genetic patterns present in the different tissues that are allowing this cool biology to occur. If we can eventually translate this knowledge to other non-symbiotic plants, it would have a massive economic impact – a Holy Grail effect."

Legumes provide about 33 percent of human nutrition in the world, but understanding and eventually being able to control nodule development in roots would benefit all plants. According to the grant's description, "the communication required to establish the symbiosis between two widely different species occurs between cell layers in tissues, between organs in the plant, and across a time span from hours to days." To understand the signaling that occurs between these cells, Frugoli and Feltus will need to determine the genes expressed in individual cells as opposed to the entire root.

"We're not only looking at what's happening in the cell. We're looking at what's happening in the cell 12 hours in, 24 hours in, 48 hours in, 72 hours in," Frugoli said. "Clemson has the ability to make RNA from very small amounts of tissue, as well as the ability to do the complex computational analysis. Oftentimes, a big project like this is spread among a number of universities. But this is entirely at Clemson because all the expertise that we need to come up with something really unique is here in one place. In addition, all our data will be freely available to scientists throughout the world. So I'm really excited about this."

Besides its research aspects, the project contains another important component: outreach. The grant provides funding to train other scientists, postdoctoral scholars and graduate students. Clemson will also bring in economically disadvantaged middle school students from around the Upstate to learn about genetics and get a feel for what it would be like to be a scientist.

"My job is to make science fun, accessible and meaningful to the students who come through our labs," said Corbin, the director of Clemson University's Life Sciences Outreach Center in the College of Agriculture, Forestry and Life Sciences. "Thirty-five to 40 middle school and high school classes come to our center each year for hands-on experiences using DNA and genetics. Unfortunately, only the wealthier school districts tend to come because they – or the parents – can pay our fees and afford buses. This grant will allow us to bring in about 100 kids from less affluent school districts each year for a lab called 'Fun with Mutants.' And we'll look at fruit fly, soil worms and also the legumes — both normal types and ones that have mutations."

Corbin said the mutations have given scientists – and hopefully will give the students – a way to determine what normal genes do for organisms, in contrast to what mutated genes do.

"It's analogous to leaving an ingredient out of a recipe. You can taste the difference between a cake with or without sugar," Corbin said. "And, of course, we will relate what we experience in the lab to human traits. Many kids see science as kind of boring, or too hard, or only for 'super smart' kids. But after they come to our outreach center, they realize that it can be fun and that they themselves can do it. That's my main mission: to inspire them to at least think about going into science and to better understand its relevance."

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The project is titled "MCR-PGR: Spatial and Temporal Resolution of mRNA Profiles During Early Nodule Development." This material is based upon work supported by the National Science Foundation (NSF) under Grant No. 1444461. Any opinions, findings and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of NSF.